Complex Vertebral Malformation (CVM) is a congenital vertebral disorder detected in Holstein-Friesian (HF) black and white dairy cattle. The disease has recently been described (Agerholm et al., 2000). In Denmark, all cases diagnosed until today (Oct. 17, 2000) have been genetically related to the former elite US Holstein bull Carlin-M Ivanhoe Bell. According to the present data, CVM appears to be inherited as an autosomal recessive disease.
The disease is characterised by a congenital bilateral symmetric arthrogryposis of the distal joints and malformations of the columna, mainly at the cervico-thoracic junction combined with reduced body weight (Agerholm et al., 1994).
Externally, there are the following major findings: In many cases the cervical and/or the thoracic part of the columna seems to be short. Moderate bilateral symmetric contraction of the carpal joints and severe contraction and supination of the phalango-metacarpal joint (fetlock) are constant findings. Contraction and pronation of the phalango-metatarsal joint and slight extension of the tarsus are also common findings. In most cases an irregular course of the columna around the cervico-thoracic junction is observed. Scoliosis may be observed, and lesions may be present in other regions of the columna. The irregular course is often recognised by inspection and palpation of the ventral aspect of the columna. However, lesions may be minimal and restricted to two or few vertebrae. In such cases the columna may be of almost normal length. Therefore, radiological examination of the columna is recommended to exclude vertebral malformations in suspected cases. The spinal cord is of normal size lying with the vertebral canal without obvious compressions. Using radiology, complex vertebral malformations consisting of hemivertebrae, fused and malshaped vertebrae, scoliosis, and anchylosis are found at varying degrees. This is best demonstrated following removal of the arcus vertebrae. In some cases malformations of the heart are present, mostly as a high interventricular septal defect and eccentric hypertrophy of the right ventricle. Malformations of the large vessels may occur. In the lungs fetal atelectasis is present. Serohemorrhagic fluids are most present in the thoracic cavity. A variety of other malformations have been observed, but these are not constant or common findings. Lesions due to dystocia are often found.
Malformations have been observed both in aborted fetuses, prematurely born calves and in stillborn calves born at term. Cases among older calves have not yet been observed. In general the body weight is reduced, and the body weight is lower in premature born calves than in calves born at term.
Additionally, there seems to be an increased frequency of abortions in cows inseminated with semen of carrier bulls. At present the cause of this is unknown.
Presently, the only tool available for CVM diagnosis is patho-anatomical diagnosis based on the above described presence of bilateral symmetric arthrogryposis of the distal joints and malformations of the columna, mainly at the cervico-thoracic junction combined with reduced body weight. However, symmetric contractions of the limbs are common and general findings in vertebral malformations in calves. Therefore, differential diagnostic problems do exist as it is often difficult to differentiate between CVM and other malformations.
The fact that the genetic defect appears to be spread by the bull Carlin-M Ivanhoe Bell which has been used intensively all over the world makes it of significant economic importance to be able to test whether current and potential breeding bulls are carriers of the defect.
In order to obtain an estimate of the frequency of potential CVM carrier animals within the Danish cattle population, the present inventors have extracted pedigree information from the Danish national cattle database. At the time of the extraction (October 2000) there were registered 919,916 pure-bred cows and heifers, and 169,821 pure-bred bulls and male calves. Bell was found 707,915 times in the pedigrees of the cows and heifers and 161,043 times in the male pedigrees. In Tables 1 and 2 below, the number of occurrences of Bell in each generation of the pedigrees is shown.
TABLE 1Occurrence of Bell in the pedigrees of Danish Holstein cows andheifersCumulativeGenerations NRFrequencyPercentFrequency2212403.021244320246028.6223704432104345.4544747513395618.96787036273073.9706010718690.37078798360.0707915
TABLE 2Occurrence of Bell in the pedigrees of Danish Holstein bullsCumulativeGenerationFrequencyPercentFrequency24360.343632014412.52058048239451.210297454454527.71475196124557.7159974710400.61610148290.0161043
Although these numbers also include some double and triple occurrences of Bell in the pedigrees, the data clearly show that a majority of the Danish Holstein cattle are potential carriers of CVM. Clearly, the problem is immense on a global scale.
Thus, there is great demand in the cattle industry for a genetic test that permits the identification of cattle in various breeds that are potential carriers of CVM (e.g. before detectable onset of clinical symptoms).
Prior to the present invention, microsatellite mapping has not been applied to the gene causing the above complex vertebral malformations which has not been isolated or characterised. Thus, to the inventors' best knowledge, the diagnostic method according to the invention described in further detail in the following has not previously been suggested or disclosed.
Accordingly, the present invention, which comprises mapping of the disease locus for CVM, has provided a DNA test based on microsatellite markers located on bovine chromosome BTA3. The ability of the test to define the carrier status of animals descending from Bell has been confirmed which appears from the examples below.